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Biomedical subjects

G W Hoyle

Publications and source records attributed to G W Hoyle.

At least 19 recordsLinked to original sources
PubMed2004

Nerve growth factor induces increased airway inflammation via a neuropeptide-dependent mechanism in a transgenic animal model of allergic airway inflammation.

BACKGROUND: Nerve growth factor (NGF) exerts an important functional impact on the pathogenesis of allergic diseases. Data obtained in animal models of allergic bronchial asthma indicate that NGF alters sensory nerve function and promotes allergic inflammation, bronchial hyper-reactivity, and airway obstruction. OBJECTIVE: To further delineate the effects of NGF on airway inflammation, we employed a transgenic (tg) animal model of allergic inflammation and asthma. METHODS: NGF-tg mice, which overexpress NGF in Clara cells of the airways, were compared with wild-type (wt) littermates regarding their ability to mount IgE-related airway inflammatory responses. Mice were sensitized intraperitoneally to ovalbumin (OVA) and locally challenged via the airways according to established protocols. RESULTS: NGF-tg mice displayed enhanced levels of OVA-specific IgE antibody titres after repeated OVA aerosol exposure. In the airways, increased numbers of eosinophils were detected. These results were confirmed to be NGF specific, because similar results were obtained following local application of NGF into the airways of wt mice. The effect of NGF was partly mediated via neuropeptides, as treatment of OVA-sensitized NGF-tg mice with the dual neurokinin (NK) receptor NK-1/NK-2 antagonist partly prevented enhanced airway inflammation. CONCLUSION: The present data indicate an important functional role of NGF in allergic airway inflammation and point to an involvement of tachykinins as mediators of NGF effects.

Animals↗
PubMed2004

Leptin receptor expression in fetal lung increases in late gestation in the baboon: a model for human pregnancy.

Leptin produced by both adipose tissue and the placental trophoblast, has been proposed to regulate numerous aspects of human conceptus development. Although recent animal studies have suggested an additional role for the polypeptide in fetal lung maturation, no evidence has been reported in primates. Therefore, we employed the baboon (Papio sp.), a well-characterized primate model for human pregnancy, to determine the presence and ontogeny of leptin receptor in fetal lung with advancing gestation. Lungs were collected from fetal baboons, early in gestation (days 58-62, n = 4), at mid gestation (days 98-102, n = 4), and late in gestation (days 158-165, n = 4) (term 184 days). mRNA transcripts for leptin (LEP) and both long and short intracellular domain isoforms of the leptin receptor (LEP-R(L) and LEP-R(S)) were assessed by RT-PCR. leptin receptor protein was evaluated by immunoblotting and cell types expressing leptin receptor were identified in late pregnancy by immunohistochemistry. Fetal serum leptin concentrations, determined by RIA, remained relatively unchanged at 5.7 +/- 1.1 ng/ml (mean +/- s.e.m.) in mid pregnancy and 8.4 +/- 3.0 ng/ml in late pregnancy (P > 0.05). Although leptin were detectable in fetal lung, no changes in transcript abundance were apparent with advancing gestation. However, transcripts for both LEP-R(L) and LEP-R(S) receptor isoforms increased several-fold (P < 0.05) in fetal lung between mid and late gestation, while leptin receptor protein was detectable only in late pregnancy. leptin receptor was localized in distal pulmonary epithelial cells, including type II pneumocytes. In conclusion, leptin is present in the fetal baboon and its receptor is enhanced during late gestation in cells responsible for the synthesis of pulmonary surfactant. Collectively, these and past findings may suggest a modulatory role for the polypeptide in pulmonary development and/or may identify leptin receptor as a physiological marker of primate fetal lung maturity.

Animals↗
PubMed2001

Overexpression of PDGF-A in the lung epithelium of transgenic mice produces a lethal phenotype associated with hyperplasia of mesenchymal cells.

Transgenic mice expressing platelet-derived growth factor A chain (PDGF-A) in the distal lung epithelium from the surfactant protein C (SPC) promoter were generated to investigate the role of this growth factor in lung development. Expression of the SPC-PDGFA transgene resulted in an enlarged, nonfunctional lung and perinatal lethality caused by failure to initiate ventilation. Histologic analysis of embryonic day (E) 16.5 lungs revealed increased mesenchymal cells and acinar buds and decreased bronchioles and dilated airspaces in SPC-PDGFA transgenic mice. At E18.5, nontransgenic lungs exhibited lung morphology typical of the saccular stage of lung development, including dilated airspaces, thin respiratory epithelium and mesenchyme, and elastin fiber deposition in primary septa. In contrast, E18.5 transgenic lungs retained many features of the canalicular stage of lung development, including undilated airspaces, cuboidal respiratory epithelium, thickened mesenchyme, and lack of parenchymal elastin deposition. These results indicate that PDGF-A is a potent growth factor for mesenchymal cells in the developing lung and that the downregulation of PDGF-A expression that normally occurs in the lung during late gestation is required for transition from the canalicular to the saccular stage of lung development.

Animals↗
PubMed2001

Sensory nerves promote ozone-induced lung inflammation in mice.

Genetically manipulated mice exhibiting altered innervation of the airways were used to examine the role of sensory nerves in ozone-induced lung inflammation. Transgenic mice expressing nerve growth factor (NGF) from the lung-specific Clara cell secretory protein (CCSP) promoter exhibit hyperinnervation of the airways by sympathetic and tachykinin-containing sensory nerve fibers. Mice carrying a mutation in the low-affinity NGF receptor (NGFR) gene possess deficits in sensory innervation. CCSP-NGF transgenic mice exhibited a twofold increase in the number of lung lavage neutrophil level whereas NGFR knockout mice exhibited a nearly 50% decrease in neutrophilic inflammation compared with wild-type mice 18 h after ozone inhalation. Treatment with neurokinin receptor antagonists reduced the level of neutrophilic inflammation in both wild-type and CCSP-NGF mice. Examination of lavage fluid cytokine concentrations revealed that 4 h after ozone exposure CCSP-NGF mice produced significantly higher amounts of the chemokine KC than wild-type mice exposed to ozone. The results of this study indicate that sensory nerves are important mediators of ozone-induced inflammation in mice.

Animals↗
PubMed1999

Reduced tumor necrosis factor-alpha and transforming growth factor-beta1 expression in the lungs of inbred mice that fail to develop fibroproliferative lesions consequent to asbestos exposure.

Tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta mRNA and protein expression and the degree of fibroproliferative response to inhaled asbestos fibers are clearly reduced in the 129 inbred mouse strain as compared with typical fibrogenesis observed in the C57BL/6 inbred strain. The C57BL/6 mice showed prominent lesions at bronchiolar-alveolar duct (BAD) junctions where asbestos fibers deposit and responding macrophages accumulate. The 129 mice, however, were generally indistinguishable from controls even though the numbers of asbestos fibers deposited in the lungs of all exposed animals were the same. Quantitative morphometry of H&E-stained lung sections comparing the C57BL/6 and 129 mice showed significantly less mean cross-sectional area of the BAD junctions in the 129 animals, apparent at both 48 hours and 4 weeks after exposure. In addition, fewer macrophages had accumulated at these sites in the 129 mice. Nuclear bromodeoxyuridine immunostaining demonstrated that the number of proliferating cells at first alveolar duct bifurcations and in adjacent terminal bronchioles was significantly reduced in the 129 strain compared with C57BL/6 mice at 48 hours after exposure (P < 0.01). TNF-alpha and TGF-beta1 gene expression, as measured by in situ hybridization, was reduced in the 129 mice at 48 hours after exposure, and expression of TNF-alpha and TGF-beta1 protein, as measured by immunohistochemistry, was similarly reduced or absent in the 129 animals. We postulate that the protection afforded the 129 mice is related to reduction of growth factor expression by the bronchiolar-alveolar epithelium and lung macrophages.

Animals↗
PubMed1999

Emphysematous lesions, inflammation, and fibrosis in the lungs of transgenic mice overexpressing platelet-derived growth factor.

Because of its expression pattern and its potent effects on mesenchymal cells, platelet-derived growth factor (PDGF) has been implicated as an important factor in epithelial-mesenchymal cell interactions during normal lung development and in the pathogenesis of fibrotic lung disease. To further explore the role of PDGF in these processes, we have developed transgenic mice that express the PDGF-B gene from the lung-specific surfactant protein C (SPC) promoter. Adult SPC-PDGFB transgenic mice exhibited lung pathology characterized by enlarged airspaces, inflammation, and fibrosis. Emphysematous changes frequently occurred throughout the lung, but inflammation and fibrotic lesions were usually confined to focal areas. The severity of this phenotype varied significantly among individual mice within the same SPC-PDGFB transgenic lineage. A pathology similar to that observed in adult mice was noted in lungs from transgenic mice as young as 1 week of age. Neonatal transgenic mice exhibited enlarged saccules and thickened primary septa. Results of these studies indicated that overexpression of PDGF-B induced distinct abnormalities in the developing and adult lung and led to a complex phenotype that encompassed aspects of both emphysema and fibrotic lung disease.

Age Factors↗
PubMed1998

TNF-alpha receptor knockout mice are protected from the fibroproliferative effects of inhaled asbestos fibers.

We have demonstrated that C57BL/6-129 hybrid mice with genes for both the 55kd and 75kd receptors for TNF-alpha knocked out (TNF-alphaRKO) fail to develop fibroproliferative lesions after asbestos exposure. There is good evidence that TNF-alpha plays a major role in mediating interstitial pulmonary fibrosis. Our findings support this view and we present here new data obtained by in situ hybridization showing that expression of the genes coding for transforming growth factor alpha (TGF-alpha) and platelet-derived growth factor A-chain (PDGF-A) is reduced in the TNF-alphaRKO mice compared with control animals. In accordance with this observation, data on bromodeoxyuridine (BrdU) incorporation in the lungs of the TNF-alphaRKO mice show no increases over unexposed control animals. In contrast, wild-type control mice exposed to asbestos exhibit 15- to 20-fold increases in BrdU uptake and consequently develop fibrogenic lesions. Even though the levels of TNF-alpha gene expression and protein production were increased in the asbestos-exposed TNF-alphaRKO mice, the lack of receptor signaling protected the mice from developing fibroproliferative lesions. We agree with the view that TNF-alpha is essential for the development of interstitial pulmonary fibrosis and postulate that TNF-alpha mediates its effects through activation of other growth factors such as PDGF and TGF-alpha that control cell growth and matrix production.

Animals↗
PubMed1998

Investigation of tissue preparation conditions for non-radioactive in situ hybridization: localization of transforming growth factor-alpha message in rat kidney.

A non-radioactive method of in situ hybridization was used to localize transforming growth factor-alpha mRNA in epithelial cells of collecting ducts and tubules in rat kidney tissue sections. The intensity and specificity of staining were assessed under a variety of tissue preparation conditions, including a direct comparison of paraffin against frozen sections. Under optimal conditions, both the signal strength and the cellular localization of the growth factor message were superior in paraffin sections. The staining method could also be used to localize the message in lung tissue, indicating that the procedure is generally applicable to other tissues. Our results indicate that the use of paraffin sections for nonradioactive in situ hybridization affords a number of advantages for the localization of specific messages in tissue sections.

Animals↗
PubMed1998

Connective tissue growth factor mRNA expression is upregulated in bleomycin-induced lung fibrosis.

Connective tissue growth factor (CTGF) is a newly described 38-kDa peptide mitogen for fibroblasts and a promoter of connective tissue deposition in the skin. The CTGF gene promotor contains a transforming growth factor-beta1 (TGF-beta1) response element. Because TGF-beta1 expression is upregulated in several models of fibroproliferative lung disease, we asked whether CTGF is also upregulated in a murine lung fibrosis model and whether CTGF could mediate some of the fibrogenic effects associated with TGF-beta1. A portion of the rat CTGF gene was cloned and used to show that primary isolates of both murine and human lung fibroblasts express CTGF mRNA in vitro. There was a greater than twofold increase in CTGF expression in both human and murine lung fibroblasts 2, 4, and 24 h after the addition of TGF-beta1 in vitro. A bleomycin-sensitive mouse strain (C57BL/6) and a bleomycin-resistant mouse strain (BALB/c) were given bleomycin, a known lung fibrogenic agent. CTGF mRNA expression was upregulated in the sensitive, but not in the resistant, mouse strain after administration of bleomycin. In vivo differences in the CTGF expression between the two mouse strains were not due to an inherent inability of BALB/c lung fibroblasts to respond to TGF-beta1 because fibroblasts from untreated BALB/c mouse lung upregulated their CTGF message when treated with TGF-beta1 in vitro. These data demonstrate that CTGF is expressed in lung fibroblasts and may play a role in the pathogenesis of lung fibrosis.

Animals↗
PubMed1998

Alveolar macrophage apoptosis and TNF-alpha, but not p53, expression correlate with murine response to bleomycin.

Apoptosis is considered to be a protective mechanism that limits lung injury. However, apoptosis might contribute to the inflammatory burden present in the injured lung. The exposure of mice to bleomycin (BLM) is a well-established model for the study of lung injury. BLM exposure induces DNA damage and enhances tumor necrosis factor (TNF)-alpha expression in the lung. To evaluate the importance of alveolar macrophage (AM) apoptosis in the pathogenesis of lung injury, we exposed BLM-sensitive (C57BL/6) and BLM-resistant (BALB/c) mice to BLM (120 mg/kg) and studied the induction of apoptosis [by light-microscopy changes (2, 8, 12, 24, 48, and 72 h) and annexin V uptake by flow cytometry (24 h)], the secretion of TNF-alpha (measured by ELISA), and the expression of p53 (by immunoblotting) in AM retrieved from these mice. BLM, but not vehicle, induced apoptosis in AM from both murine strains. The numbers of apoptotic AM were significantly greater (P < 0.001) in C57BL/6 mice (52.9%) compared with BALB/c mice (40.8%) as demonstrated by annexin V uptake. BLM induction of apoptosis in AM was preceded by an increased secretion of TNF-alpha in C57BL/6 but not in BALB/c mice. Furthermore, double TNF-alpha receptor-deficient mice, developed on a C57BL/6 background, demonstrated significantly (P < 0.001) lower numbers of apoptotic AM compared with C57BL/6 and BALB/c mice. BLM also enhanced p53 expression in AM from both murine strains. However, p53-deficient mice developed BLM-induced lung injury, exhibited similar lung cell proliferation (measured as proliferating cell nuclear antigen immunostaining), and accumulated similar amounts of lung hydroxyproline (65 +/- 6.9 microgram/lung) as did C57BL/6 (62 +/- 6.5 microgram/lung) mice. Therefore, AM apoptosis is occurring during BLM-induced lung injury in a manner that correlates with murine strain sensitivity to BLM. Furthermore, TNF-alpha secretion rather than p53 expression contributes to the difference in murine strain response to BLM.tumor necrosis factor; strain susceptibility

Animals↗
PubMed1998

Hyperinnervation of the airways in transgenic mice overexpressing nerve growth factor.

Neuropeptides released from sensory nerve endings are potential mediators of airway inflammation in asthma and lung injury induced by inhalation of respiratory irritants. To develop an in vivo model for assessing the contribution of neurogenic inflammation in these processes, we have generated transgenic mice with altered innervation of the lung. To generate mice with an increased innervation of the airways, we placed the gene that encodes nerve growth factor (NGF) under control of the lung-specific Clara-cell secretory protein (CCSP) promoter. Two lineages of CCSP-NGF transgenic mice overexpressed NGF in the lung and developed a hyperinnervation of the airways. Immunohistochemistry for substance P, a substance P enzyme immunoassay, and catecholamine histofluorescence indicated that both tachykinin-containing sensory fibers and sympathetic fibers were increased around the airways of CCSP-NGF mice. Treatment of CCSP-NGF mice with the sympathetic-specific neurotoxin 6-hydroxydopamine (6-OHDA) eliminated the sympathetic component of the airway innervation, leaving a specific hyperinnervation by tachykinin-containing sensory fibers. CCSP-NGF mice were more sensitive than normal mice to capsaicin-induced increases in respiratory system resistance, demonstrating that the increased sensory innervation led to a change in airway function. We conclude that NGF overexpression from a lung-specific promoter produces anatomic and functional changes in lung innervation, and that CCSP-NGF mice will be useful for studying the role of neurogenic inflammation in airway disease.

Airway Resistance↗
PubMed1998

Lung-specific expression in mice of a dominant negative mutant form of the p53 tumor suppressor protein.

Lung cancer is the most frequent cause of cancer deaths in the United States. A strong correlation exists between mutations in the gene encoding the p53 tumor suppressor protein and lung malignancies. Our goal is to prepare a transgenic mouse model with disrupted p53 function in the epithelial cells of the peripheral lung. To achieve this goal, a "dominant negative" mutant form of p53 was expressed from the human surfactant protein C (SPC) promoter. The dominant negative p53 expressed from the SPC promoter will antagonize wild-type p53 functions in alveolar type II pneumocytes and some bronchiolar cells of the transgenic animals and thereby promote development of carcinoma of the lung. This animal model should prove useful to the study of lung carcinogenesis and to the identification of agents that contribute to neoplastic conversion in the lung.

Animals↗
PubMed1994

Leukemia inhibitory factor induces neurotransmitter switching in transgenic mice.

Leukemia inhibitory factor (LIF) is a cytokine growth factor that induces rat sympathetic neurons to switch their neurotransmitter phenotype from noradrenergic to cholinergic in vitro. To test whether LIF can influence neuronal differentiation in vivo, we generated transgenic mice that expressed LIF in pancreatic islets under the control of the insulin promoter and evaluated the neurotransmitter phenotype of the pancreatic sympathetic innervation. We also used the insulin promoter to coexpress nerve growth factor in the islets, which greatly increased the density of sympathetic innervation and facilitated analysis of the effects of LIF. Our data demonstrate that tyrosine hydroxylase and catecholamines declined and choline acetyltransferase increased in response to LIF. We conclude that LIF can induce neurotransmitter switching of sympathetic neurons in vivo.

Aging↗
PubMed1994

Visualization and ablation of phenylethanolamine N-methyltransferase producing cells in transgenic mice.

We cloned and sequenced the mouse phenylethanolamine N-methyltransferase (PNMT) gene which encodes the enzyme that catalyses the conversion of norepinephrine to epinephrine. The ability of various length sequences flanking the mouse or human PNMT genes to direct expression of reporter genes in transgenic mice was examined. We show that 9 kb of 5' flanking sequences from the cloned mouse PNMT gene can direct expression of the Escherichia coli beta-galactosidase (lacZ) gene to predicted regions of the adrenal, eye and brain in the adult transgenic mouse. The transgene was also expressed during development, in the myelencephalon, adrenal medulla and dorsal root ganglia. PNMT-producing cells were ablated by expression of the diphtheria toxin (DT-A) gene driven by the human PNMT promoter, resulting in abnormalities in the adrenal medulla, eye and testis. The hPNMT8 kb-DT-A line presents a model with which to examine the developmental ramifications of deletion of PNMT-producing cell populations from the adrenal medulla and retina.

Adrenal Medulla↗
PubMed1994

Cell-specific expression from the human dopamine beta-hydroxylase promoter in transgenic mice is controlled via a combination of positive and negative regulatory elements.

The promoter region of the human dopamine beta-hydroxylase (DBH) gene was analyzed in transgenic mice to identify DNA sequences responsible for the tissue- and cell-specific expression of the gene. Transgenic mice were generated that carried the Escherichia coli lacZ gene under control of DBH promoter fragments between 0.6 and 5.8 kilobases (kb) in length. Sequences required for expression in adult and fetal noradrenergic neurons were located between 0.6 and 1.1 kb 5' to the DBH transcriptional start site. Sequences in this region and farther upstream also directed expression to dopaminergic and noncatecholaminergic brain neurons that was repressed by negative elements elsewhere in the gene. The results indicate that the neuron-specific expression of the DBH gene is mediated by positive regulatory elements but that negative elements are required to restrict expression to the proper subset of neurons.

Animals↗